Analysis of Retinoblastoma Transcriptome
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE141209 | GEO | 2020/06/02
REPOSITORIES: GEO
Similar Datasets
Project description:Background: Retinoblastoma (RB) is the most common malignant childhood tumor of the eye and results from inactivation of both alleles of the RB1 gene. Nowadays RB genetic diagnosis requires classical chromosome investigations, Multiplex Ligation-dependent Probe Amplification analysis (MLPA) and Sanger sequencing. Nevertheless, these techniques show some limitations. We report our experience on a cohort of RB patients using a combined approach of Next-Generation Sequencing (NGS) and RB1 custom array-Comparative Genomic Hybridization (aCGH). Methods: A total of 65 patients with retinoblastoma were studied: 29 cases of bilateral RB and 36 cases of unilateral RB. All patients were previously tested with conventional cytogenetics and MLPA techniques. Fifty-three samples were then analysed using NGS. Eleven cases were analysed by RB1 custom aCGH. One last case was studied only by classic cytogenetics. Finally, it has been tested, in a lab sensitivity assay, the capability of NGS to detect artificial mosaicism series in previously recognized samples prepared at 3 different mosaicism frequencies: 10%, 5%, 1%. Results: Of the 29 cases of bilateral RB, 28 resulted positive (96.5%) to the genetic investigation: 22 point mutations and 6 genomic rearrangements (four intragenic and two macrodeletion). A novel germline intragenic duplication, from exon18 to exon 23, was identified in a proband with bilateral RB. Of the 36 available cases of unilateral RB, 8 patients resulted positive (22%) to the genetic investigation: 3 patients showed point mutations while 5 carried large deletion. Finally, we successfully validated, in a lab sensitivity assay, the capability of NGS to accurately measure level of artificial mosaicism down to 1%. Conclusions: NGS and RB1-custom aCGH have demonstrated to be an effective combined approach in order to optimize the overall diagnostic procedures of RB. Custom aCGH is able to accurately detect genomic rearrangements allowing the characterization of their extension. NGS is extremely accurate in detecting single nucleotide variants, relatively simple to perform, cost savings and efficient and has confirmed a high sensitivity and accuracy in identifying low levels of artificial mosaicisms.
2015-10-31 | E-MTAB-3492 | biostudies-arrayexpress
Project description:Bilateral retinoblastoma has served as a critical model for studying genetic predisposition to cancer, however international incidence of unilateral retinoblastoma is variable across populations suggesting the involvement of environmental factors. DNA from Human Papilloma Virus (HPV) has been found in human retinoblastoma tissue through polymerase chain reaction based methods in several independent reports. To search for HPV oncoproteins and characterize transcriptional profiles in retinoblastoma, we analyzed tumor tissue obtained after enucleation and cultured for one week from 13 previously untreated Mexican children with retinoblastoma. We identified the presence of E7 HPV oncoproteins using western blot analysis in HPV positive tumors, and using non-supervised methods of classification on gene expression microarrays data, tumors clustered according to the presence or absence of HPV DNA identified by PCR. These methods were unable to discriminate retinoblastoma tissues by laterality. 2,404 genes were differentially expressed between HPV positive and HPV negative retinoblastoma, including a group of genes involved in a transcriptional response to viral infection. This molecular evidence supports the hypothesis that HPV is involved in the molecular pathogenesis of these tumors. The 13 children and tumors examined here, are incident cases participating in a larger case-control study at two Pediatric Hospitals of third level of medical attention in Mexico City, that form part of the public health system. Nine different tumors positive and four negative to HPV by PCR were included in the experimental design. From these, seven are unilateral and six are bilateral and are considered biological replicas. Two unilateral and two bilateral cases were chosen to make technical replicas. From the surgical specimens, two tumor portions were obtained, one was frozen, DNA was extracted from it, and HPV status was assigned using MY09 and MY11 primers by PCR. Another portion of tumor tissue was used to grow in culture, separating viable cells from non viable cells through a ficoll gradient. In the eye, retinoblastoma usually grows into the vitreous which has no cells, and thus the tissue obtained is predominantly formed by tumor cells and few endothelial cells from capillaries in the tumor. Retinoblastoma grow in culture as non adherent cells, and when adherent cells were also present, cell suspension was moved to a new dish to keep retinoblastoma cells as the main or only cellular type as possible. We used a two channel platform with in house spotted microarrays. Since retinoblastoma has known classes: uni and bilateral, we sought to verify these classes or discover new, we used a class discovery approach and a reference experimental design. As the reference sample we used RNA form exponentially growing Saccharomyces cerevisiae because major signaling pathways and basic cellular processes among Saccharomyces cerevisiae and mammalian cells including cell cycle control and DNA repair mechanisms have a high degree of conservation. Also because yeast is easy to grow, and thus a reliable and robust source of reference RNA.
2008-05-20 | E-GEOD-11488 | biostudies-arrayexpress
Project description:Bilateral retinoblastoma has served as a critical model for studying genetic predisposition to cancer, however international incidence of unilateral retinoblastoma is variable across populations suggesting the involvement of environmental factors. DNA from Human Papilloma Virus (HPV) has been found in human retinoblastoma tissue through polymerase chain reaction based methods in several independent reports. To search for HPV oncoproteins and characterize transcriptional profiles in retinoblastoma, we analyzed tumor tissue obtained after enucleation and cultured for one week from 13 previously untreated Mexican children with retinoblastoma. We identified the presence of E7 HPV oncoproteins using western blot analysis in HPV positive tumors, and using non-supervised methods of classification on gene expression microarrays data, tumors clustered according to the presence or absence of HPV DNA identified by PCR. These methods were unable to discriminate retinoblastoma tissues by laterality. 2,404 genes were differentially expressed between HPV positive and HPV negative retinoblastoma, including a group of genes involved in a transcriptional response to viral infection. This molecular evidence supports the hypothesis that HPV is involved in the molecular pathogenesis of these tumors.
2008-05-21 | GSE11488 | GEO
Project description:Retinoblastoma (Rb) is the most common intraocular tumor in childhood and represent a very robust model about hereditary develop for cancer. We believe that miRNome expression may give clues to understanding Rb heterogeneity and to comprehend biological features in order to find new biomarkers for better treatments. We explore 12 Rb tumors from enucleated patients with no chemotherapy history from Children Hospital of Mexico Federico Gomez and Pediatric Hospital Medical Center XXI century by informed parental consent to collaborate with retrospective research. 5 bilateral and 7 unilateral, 4 female and 8 male samples were tailed and labeled with biotin to hybridize in GeneChip miRNA 4.0 Affymetrix array. Chips were washed and scanned 18hrs after hybridization.
2017-07-11 | E-MTAB-4977 | biostudies-arrayexpress
Project description:Transcriptome analysis of partially degraded and fragmented RNA samples from retinoblastoma primary tumors. Global gene expression profiling has shown great promise in high-throughput biomarker discovery for early disease detection in Retinoblastoma tumors. In this analysis we have used 3 retinoblastoma primary tumors in triplicates which was normalised against human healthy Retina. Here, we have used nanogram scale of retinoblastoma RNA, processed in HUMAN GENE 1.0 ST ARRAYS.
2010-10-14 | GSE24673 | GEO
Project description:Transcriptome analysis of partially degraded and fragmented RNA samples from retinoblastoma primary tumors. Global gene expression profiling has shown great promise in high-throughput biomarker discovery for early disease detection in Retinoblastoma tumors. In this analysis we have used 3 retinoblastoma primary tumors in triplicates which was normalised against human healthy Retina. Here, we have used nanogram scale of retinoblastoma RNA, processed in HUMAN GENE 1.0 ST ARRAYS. We have analysed the gene expression in 2 normal healthy adult retina collected from cadaveric eyes and 3 retinoblastoma primary tumors
2010-10-14 | E-GEOD-24673 | biostudies-arrayexpress
Project description:mRNA-seq of retinoblastoma samples
| PRJEB44185 | ENA
Project description:Retinoblastoma (RB, OMIM:180200) is the most common malignant childhood tumor of the eye with an estimated incidence between 1 in 16,000 and 1 in 18,000 live births [1,2]. RB is the first disease for which a genetic etiology of cancer has been described [3] being caused by mutations in the first tumor suppressor gene identified (RB1, Genbank accession # L11910). Mutations in both alleles of the RB1 gene are required for the development of this neoplasm [4], and, depending on the germ-line or somatic origin of the defect, a heritable or sporadic form can be distinguished. RB is unilateral in 60% of cases and only 15% of these are heritable [5]; in contrast, 40% of retinoblastomas are bilateral with risk of transmission to the offspring. Heritable retinoblastoma constitutes a cancer predisposition syndrome [6]. RB1 is located on chromosome 13 at band q14 and can be affected by a heterogeneous spectrum of genetic abnormalities, including chromosome translocation/deletion, genomic rearrangements, ranging from whole gene microdeletion to intragenic exons loss or duplication, and more than 900 different point mutations [7]. Mutational analysis is performed to search for the predisposing RB1 gene mutation in peripheral blood of patients with RB, but the molecular diagnosis requires several technical approaches to cover the entire field of oncogenic RB1 defects, frequently resulting in numerous, expensive and time consuming procedures. In particular, cytogenetic tools, such as classical chromosome investigations and Fluorescent In Situ Hybridization (FISH), in addition to Multiplex Ligation-dependent Probe Amplification (MLPA) technique, may account for detection of about 16% of RB1 abnormalities [8], while the remaining large amount of point mutations need to be investigated using sequencing analysis. Since the 1970s, Sanger sequencing has been recognized as the gold standard for mutation analysis in molecular diagnostics; however, its low-throughput, long turnaround time and overall cost [9] have called for new paradigms. Next Generation Sequencing (NGS) can massively sequence millions of DNA segments, promising low costs, increased workflow speed and enhanced sensitivity in mutation detection [9,10,11] On the other hand, conventional and molecular cytogenetic analysis, have been replaced by modern high-throughput investigations, such as array Comparative Genomic Hybridization (aCGH), that can reveal and measure cryptic genomic imbalances. In addition, aCGH can be focused on specific DNA segments or genes maximizing the resolution via a customized process. Based on these observations, we have recruited a cohort of retinoblastoma patients we previously investigated with conventional cytogenetics and MLPA. Patients diagnosed with RB but negative to the above standard screening have been tested with NGS to assess its ability in identifying RB causative mutations. On the other hand, patients positive to standard screening have been further investigated with RB1-custom array CGH analysis to characterize the genomic rearrangements with a better resolution compared to the conventional techniques. The complementary aCGH data set related to this study has also been deposited at ArrayExpress, under accession number E-MTAB-3492: https://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-3492/
2015-10-31 | E-MTAB-3515 | biostudies-arrayexpress
Project description:In order to identify the gene targets of frequently altered chromosomal regions in retinoblastoma, a meta-analysis of genome-wide copy number alterations studies on primary retinoblastoma tissue and retinoblastoma cell lines was performed. Published studies were complemented by copy number and gene expression analysis on primary and cell line samples of retinoblastoma. This dataset includes the gene expression data of the retinoblastoma cell lines
2016-03-31 | GSE77094 | GEO
Project description:To obtain a global view of APA during the development of retinoblastoma, we subjected total RNAs from five retina sample and five retinoblastoma samples to PA-seq, which precisely identify poly(A) sites at the genome scale.
2023-02-03 | GSE224022 | GEO